The broad, long-term objectives and specific aims of the proposed research are the elucidation of salt and water transport by the kidney. It is planned to study the role of cell calcium ([cA2+]I) and protein kinase C (PKC)-mediated regulation of the amilordie- sensitive, small conductance Na channel. This channel is under the control of aldosterone and vasopressin and plays a critical role in the maintenance of salt and water balance in the body. More specifically, in rat renal cortical collecting tubules (CCTs), the effect of EGF will be assessed.- To test whether stimulation by cAMP masks the inhibitory action of high [Ca2+]I on Na channel activity the effect of ionomycin on channel activity will be examined in cAMP-pretreated CCTs. Furthermore, the identity of the specific isoforms of PKC in principal cells of the CCT and the degree to which they depend on changes in [Ca2+]I for activation and translocation will be determined by immunofluorescence imaging of antibodies specific to the isoforms.- using the whole cell patch clamp approach we will study the effect of specific inhibitors of PKC and Ca2+/CaM kinase II and of the addition of activated forms of these kinases on the Ca2+-mediated inhibition of the amiloride-sensitive Na current in principal cells. With the same approach, the effect of dephosphorylation by protein phosphatases will be explored.- Finally, the role of Na-self-inhibition of Na channels in the regulation of apical Na permeability in principal cells will be explored.